Cut It Out
Gene therapy can be used against viruses as well. Cellectis Therapeutics (a subsidiary of Cellectis) uses meganucleases engineered to precisely disrupt integrated viruses such as HIV, HBV, and HSV.
Meganucleases (also known as homing endonucleases) target a specific, statistically unique, sequence—generally at least 14 base pairs—creating a double-strand break in the DNA. The cell then attempts to repair the break, often with some minor mutation or loss of sequence.
Cellectis’ most advanced meganuclease project targets an essential gene in the herpes simplex virus. “We were able to show that if we take cells and introduce a plasmid that expresses a meganuclease that targets the viral genome, and then we infect those cells with an HSV virus, we can decrease the infectivity of that virus,” related Julianne Smith, Ph.D., head of the meganuclease recombination system group. “The number of infected cells significantly decreases when the meganuclease is present.”
If there is a homologous sequence present, the cell may attempt to repair the break by homologous recombination. In a therapeutic context, “you could go in and actually create a double-stranded break and introduce a plasmid that contains wild-type information, and go on to correct the mutated allele at its genomic location.”
To test this idea, Cellectis developed a series of meganucleases targeting the RAG-1 gene (involved in a form of severe combined immunodeficiency), and used them to demonstrate homologous recombination between the chromosomal locus and a repair plasmid.
The efficiency by which meganucleases get into cells and do their job is about the same as other means of gene therapy. The important difference is the safety aspect, said Dr. Smith. “If we can specifically target a genomic site, we could avoid some of the serious adverse effects that have been observed concerning random insertions and activation of adjacent genes.” Similarly, random insertions are also subject to gene silencing through gene extinction, but with precise targeting you can select your locus.
In certain cases, it’s even possible to use meganucleases to generate an event in a cell that couldn’t be done any other way, she added, such as cleaving the genome of a virus or perhaps even knocking out a gene. Most antivirals are based on blocking replication, not on eliminating the genome.